The blood-brain barrier (BBB) separates the blood and the central nervous system (CNS). The central nervous system consists of the brain and the spinal cord. This separation between the blood and CNS is created by specialized endothelial cells, brain endothelial cells, which are distinct from peripheral endothelial cells, as the blood-brain barrier is composed of high density cells restricting passage of substances from the bloodstream much more than endothelial cells elsewhere in the body (Abbott et al., Nat Rev Neurosc, 2006, 7, 41-53). The BBB restricts the diffusion of large or hydrophilic molecules into the CNS, e.g. antibodies, but also of microscopic entities, e.g. bacteria. On the other hand, small hydrophobic molecules, e.g. hormones and oxygen, can diffuse over the BBB. The BBB also actively transports molecules, such as for instance glucose, across the BBB utilizing transporter proteins. Cells of the immune system can also pass the BBB (Pachter et al., J Neuropathol Exp Neurol. 2003 June; 62(6):593-604; Abbott et al., Nat Rev Neurosc, 2006, 7, 41-53; Begley and Brightman, Prog Drug Res. 2003; 61:39-78; Loscher and Potschka, NeuroRx. 2005 January; 2(1):86-98; Carvey et al., Neurochem. 2009 October; 111(2):291-314; Abbott et al., Neurobiol Dis. 2010 January; 37(1):13-25). In summary, the function of the BBB is to allow precise control over the substances that leave or enter the central nervous system, and this function is essential for brain homeostasis and reliable functioning of the neuronal environment.
A common feature of diverse brain abnormalities such as for example multiple sclerosis, brain cancer, Alzheimer's disease, stroke, epilepsy and traumatic brain injury, is reduction or loss of the specialized function of the BBB, leading to unstable brain homeostasis and neuronal damage (Zlokovic, Neuron. 2008 Jan. 24; 57(2):178-201; Friedman et al., Epilepsy Res. 2009 August; 85(2-3):142-9; Carvey et al., Neurochem. 2009 October; 111(2):291-314; Abbott et al., Neurobiol Dis. 2010 January; 37(1):13-25).
On the other hand, in brain abnormalities in which the BBB function is maintained, the BBB can obstruct or hamper the entry of pharmaceuticals or drugs. For instance, many high potential drugs for the central nervous system (CNS), for instance biopharmaceuticals such as antibodies, are currently not or less suitable for use in the treatment of the CNS because these drugs can often not cross the blood-brain barrier, making the BBB a major hurdle for successful CNS drug development that require drugs to pass the BBB.
Current strategies for drug delivery in the brain involve invasive measures such as for instance stereotactic injections in the brain or intrathecal injections, which both means the BBB is physically disrupted and which always involves a risk of neural injuries. Other means include disruption of the BBB by osmotic means, or biochemically by the use of vasoactive substances such as bradykinin or even by localized exposure to high intensity focused ultrasound (HIFU), but success has been limited (Daffertshofer and Fatar, Eur J. Ultrasound. 2002 November; 16(1-2):121-30; Doolittle et al., J Neurosci Nurs. 1998 April; 30(2):81-90; Wahl et al., Acta Physiol Hung. 1999; 86(2):155-60). As such, there remains a need for methods and compounds that can safely and effectively modulate, i.e. increase or decrease the blood brain barrier function. With an increase of the blood brain barrier function is meant that the blood brain barrier becomes less permeable for the passive transfer of compounds across the BBB, for example (therapeutic) antibodies or antibiotics. With a decrease of the blood brain barrier function is meant that the blood brain barrier becomes more permeable for the passive transfer of compounds such as for example (therapeutic) antibodies or antibiotics across the BBB. Such compounds or methods that would influence the BBB function would be beneficial, as for instance co-administration of a compound that decreases the blood brain barrier with a drug, e.g. therapeutic antibodies or antibiotics, that normally is less capable or incapable of passing the BBB might result in the improved delivery of said drug to the brain. In addition, a compound that increases the blood brain barrier function may allow the (partial) reconstitution of the blood brain barrier function in patients that suffer from impaired BBB function. An increase of the BBB function might for instance prevent bacterial infiltration of the brain or reduce seizures, or help to restore homeostasis.
Unfortunately, the discovery of compounds that may modulate the BBB has been hampered by a lack of understanding of the functioning of the BBB and the pathways involved, as well by the absence of easy methods to determine BBB function. For instance, in vivo animal knockout models were used in which the involvement of certain transmembrane proteins involved in the BBB function such as for instance the G protein-coupled receptor 124 was observed (GPR124) (WO2008147528). Downregulating GPR124 with an antagonist was suggested to decrease the BBB function. On the other hand, increasing expression of GPR124 may provide for increased BBB function. In addition, it has also been suggested that inhibiting claudin-5 expression using RNA interference with hydrodynamic tail vein injections may be used to “open up” the BBB (WO2009047362). In such animal experiments the permeability of the BBB is determined by measuring the uptake by the brain of tracer molecules that normally have difficulty or can not pass the BBB. Obviously, in vivo animal models or knock-out studies do not allow large scale screening for new candidate compounds.
A less complicated assay to study the BBB function is an in vitro assay in which (primary) brain endothelial cells are cultured in vitro on a membrane and passage of tracer compounds through the membrane is determined. Alternatively, the electrical resistance (expressed as ohm) over the cultured brain endothelial cells can also be measured, i.e. the transendothelial electric resistance (Deli et al., Cell Mol. Neurobiol. 2005 February; 25(1):59-127). Importantly, in many of these in vitro assays the cells have to be grown to full confluency in order to ensure that the tracer compounds (or current) has to pass the brain endothelial cell layer and does not simply bypass the membrane without encountering the brain endothelial cells. Although these in vitro models are a major improvement over the in vivo experiments, they are laborious and technically challenging, making large scale screening to identify compounds that influence BBB function still a very difficult task.
In summary, a problem in the development of a medical treatment for the central nervous system, e.g. the brain, is that there is a need for compounds that can safely and effectively modulate, i.e. increase or decrease, the blood brain barrier function. In addition, there is a need for assays that are robust and simple to perform to allow for large scale screening for compounds (or compound libraries) that can modulate the blood brain barrier function.